真菌Polyporus sp. M05多糖诱导白血病细胞株HL-60凋亡作用机制的实验研究

Effect and mechanism of apoptosis HL-60 cells induced by a fungus polysaccharide

目的:研究真菌Polyporus sp.M05多糖对白血病细胞株HL-60增殖抑制及诱导凋亡作用,并探讨其分子机制。方法:活细胞计数法检测多糖的增殖抑制作用;倒置显微镜下直接观察细胞形态;PI染色流式细胞术检测多糖诱导周期阻滞及凋亡作用;RT-PCR检测凋亡相关基因。结果:多糖对HL-60细胞有增殖抑制作用,并随着浓度增大,作用增强;显微镜下观察细胞有明显的凋亡小体。流式细胞术检测细胞有明显G0/G1期阻滞及凋亡峰的出现,并随着时间越长和浓度的增大凋亡阳性率比例越高,与阴性对照相比差异有统计学意义(P<0.01);RT-PCR检测到Bcl-2基因下调,Bax、Fas及Caspase-9基因上调。结论:Polyporus sp.M05中提取的多糖对白血病细胞株HL-60有增殖抑制和诱导凋亡作用,分子机制与Bcl-2基因下调,Bax、Fas及Caspase-9基因上调有关,线粒体途径及死亡受体途径均参与了此凋亡过程。

OBJECTIVE： To study the effect and mechanism of apoptosis of HL-60 cells induced by polysaccharide PSM from Polyporus sp.M05.METHODS： The proliferation and inhibition were studied by the method of leukocyte counting.The cell morphology was detected through a inverted microscope.The cell cycle and apoptosis were analyzed by flow cytometry with propidium iodide staining.Semi-quantitative RT-PCR was used to detect the mRNA expression of the apoptosis gene.RESULTS： PSM significantly inhibited the proliferation of HL-60 cells.The apoptosis bodies were obviously detected through the inverted microscope.After being induced by PSM,the cell cycle was significantly arrested at the G0/G1 phase.The apoptosis ratio was increased significantly in a time and dose manner.After being induced by PSM,mRNA of Bcl-2 was down regulated and mRNA（s） of Bax,Fas,Caspase were up regulated.CONCLUSIONS： The mechanism of HL-60 cell apoptosis induced by PSM is related to the up-regulation of Bax,Fas,Caspase and down-regulation of Bcl-2,and involved in the death receptor pathway and mitochondia pathway.